Inflammatory response of cultured rat synoviocytes challenged with synovial fluid from osteoarthritis patients correlates with their radiographic grading: a pilot study

Abstract

The inflammatory nature of synovial fluid (SF) of varying grade osteoarthritis (OA) patients was estimated by measuring pro-inflammatory factors and through a unique cell-challenge experiment. SF samples were collected from six OA and one non-OA patient; spanning Kellgren-Lawrence (KL) grades were analyzed for interlukin-1-beta (IL-1β), nitric oxide (NO) and its derivatives, and glycosaminoglycan (GAG). Levels of IL-1β, NO, and GAG in SF did not correlate with KL grades of the patients studied. In the cell-challenge experiment, cultured rat synoviocyte fibroblasts (RSFs) were challenged by the patient's SFs with and without pre-treatment of IL-1β and lipopolysaccharide (LPS). NO released by the cells was taken as an indicator of inflammation. SFs from KL grades 2 and 3 induced maximum inflammation in cultured RSFs (grade 2 64.61 ± 4.8 and 89.51 ± 5.6 μM/ml after 48 and 72 h, grade 3 58.27 ± 2.7 and 64.22 ± 2.8 μM/ml after 48 and 72 h, respectively). Similar trend was observed in RSF pretreated with either recombinant IL-1β or LPS suggesting that SF from patients KL grades 2 and 3 accumulates more pro-inflammatory factors. IL-1β-pre-treated RSFs challenged by SF for 72 h showed 234.41 ± 17.6 μM/ml increase (patient 3, grade 3), whereas higher NO after LPS pre-treatment was recorded (118.92 ± 6.2 μM/ml; patient 3, grade 3). Interestingly, SFs from grade 1 and non-OA patient could reduce released NO to 27.10 ± 2.2 μM/ml showing potency to alleviate inflammation. These interesting findings, however, need to be confirmed on a wider number of patients, which may offer significant therapeutic application in treatment of OA.

Synovium-Synovial Fluid Axis in Osteoarthritis Pathology: A Key Regulator of the Cartilage Degradation Process

“…The novelty of the present data is the SF challenge test that was designed to study the dynamics of modulation in inflammatory microenvironment at joint in terms of classical inflammatory mediators and MMPs expression. These experiments were based on our previous learning that OA SF holds inflammatory milieu of various cytokines and chemokines and can be used to study the disease specific responses on cell lines [ 14 ]. Most of the etiopathology of OA spins around the synovium-SF-cartilage axis, wherein SF occupies the central position.…”

“…IL-1β expression in the SF treated SW982 was tested by qPCR and the values were normalized against β-actin gene. SF challenge test was developed and standardized by our research group and was successfully used to study OA specific responses [ 14 ].…”

Mast Cells Differentiated in Synovial Fluid and Resident in Osteophytes Exalt the Inflammatory Pathology of Osteoarthritis

“…Lastly, the maximum differentiation of ThP1 and HSCs into the effector cells was seen after the SF treatment of KL grade III. This is likely to be the effect of the highest inflammatory milieu present in the KL grade III SF samples as reported by the authors in their previous publications [ 40 ]. On the other hand, this inflammatory milieu was found reduced in the KL grade IV SF samples as an effect of near-total worn-out cartilage in advanced OA [ 41 ] and is a possible reason for the relatively lower number of differentiated cells after the SF treatment of KL grade IV samples.…”

“…The hypothesis of the in vitro cell differentiation assay was to test the role of OA SF in the maturation of MCPs by providing the necessary cocktail of cytokines, cell degradation products, and immunoglobulins. In the previous studies, the authors showed that this cocktail can be used to induce inflammation in SW982, U937, HIG82 and ThP1 cell lines [ 1 , 40 , 41 ]. As demonstrated by these assays, the differentiation of HSCs and ThP1 cells into FCERI+ and HLA-DR+/CD206+ phenotypes, respectively, after SF treatment, indicate a decisive role of OA SF in the differentiation of polarised cells into effector cells such as mast cells and macrophages ( Figure 5 a–c).…”

Mast cells differentiated in synovial fluid and resident in osteophytes exalt the inflammatory pathology of osteoarthritis

“…OA SF plays a major role in maturation of MCPs by providing necessary cocktail of cytokines, cell degradation products, and immunoglobulins. In our previous studies, we showed that this cocktail can be used to induce inflammation in HIG82, SW982, ThP1 and U937 cell lines [13,39,40]. As demonstrated by in vitro cell differentiation assays, differentiation of HSCs and ThP1 cells into FCERI+ and HLA-DR+/CD206+ phenotypes, respectively after SF treatment, indicate a decisive role of OA SF in differentiation of polarised cells into effector cells such as mast cells and macrophages (Figure 4 -a, b, c).…”